1. Field of the Invention
The present invention relates generally to inflammation and, more particularly, to treating inflammation, regulating cytokines, and kits therefor.
2. Description of the Prior Art
Inflammation is a normal part of the response to injuries, invasion by pathogens, and may occur without known cause. The inflammatory process can protect an organism by eliminating pathogens or by removing injured tissue and promoting the restoration of new tissue. However, overabundant or persistent inflammation results in the malfunction or the destruction of vital cells and tissues. Dysregulated inflammation is a hallmark of many painful and life threatening diseases and can affect every tissue and organ of the body.
Persistent or chronic inflammation is often characterized by increased production of pro-inflammatory mediators including cytokines, such as, interleukin 6 (IL-6) and interleukin 8 (IL-8). The stimulated expression of IL-6 and IL-8 is thought to be regulated primarily through increased transcription. Therefore, the promoter regions of IL-6 and IL-8 have been examined to determine which transcription factors activate expression. Two transcription factors are identified as being critical for maximal IL-6 and IL-8 expression: nuclear factor-κB (NF-κB) and CCAAT/Enhancer Binding Protein (C/EBP). C/EBP is actually a group or family of transcription factors related by sequence, structure, and/or biological activity. It has been demonstrated that one member of the C/EBP family, C/EBPβ, can form a complex with NF-κB and DNA motifs on the promoter(s) of a target gene leading to a synergistic activation of transcription (including the IL-1β, IL-6, and IL-8 genes among others, see, e.g., Stein et al., Molecular Cell Biol. (1993) 13:3964–3974; Lee et al., Molecular Cell Biol. (1996) 16:4257–4263; Kunsch et al., (1994) J. Immunol. 153:153–164; and Poli (1998) J Biol. Chem. 273(45):29279–29282).
C/EBPβ and Isoforms thereof
In human, murine, and rat cells, three isoforms of C/EBPβ are observed and are referred to as C/EBPβ-1, C/EBPβ-2, and C/EBPβ-3 (in order from longest to shortest). The three isoforms correlate to three in frame open reading frames (ORFs) in the C/EBPβ gene. The shortest C/EBPβ isoform, C/EBPβ-3, is referred to herein as p20 (a protein of approximately 20 kDa molecular weight).
The transcription of a recombinant C/EBPβ responsive reporter gene was found to be activated by transfection of a C/EBPβ-2 expressing gene and inhibited by co-transfection of a p20 gene in cultured cells (Descombes et al. (1991) Cell 67:569–579).
U.S. Pat. No. 5,804,445 describes an isolated 8.8 kDa tryptic fragment of C/EBPβ with mutations in the N-terminus of the fragment which make the N-terminus of the fragment less negative. The mutated tryptic fragment has a higher binding affinity for purified DNA containing a C/EBPβ binding site compared to similar fragments with the wild-type sequence.
U.S. Pat. No. 5,874,209 describes a peptide consisting of amino acids 75 to 125 of C/EBPβ, wherein residue 105 is mutated from serine to alanine to prevent phosphorylation of the 105 residue. The 105 mutated peptide competes with native C/EBPβ in cultured cells to inhibit transactivation of a reporter gene.
The Effects of Dysregulated Inflammation
Virtually all diseases of the lungs have an inflammatory component. Disorders such as idiopathic pulmonary fibrosis (IPF), adult respiratory distress syndrome (ARDS), cystic fibrosis (CF) and asthma, in particular, are characterized by over exuberant or persistent lung inflammation. In addition, overabundant or chronic inflammation is a component of many other diseases and often leads to the destruction of vital cells, tissues, and organs (e.g., the digestive tract organs and the heart and blood vessels). Currently, nonspecific suppression of inflammation with high doses of corticosteroids is used to treat these disorders. However, high dose corticosteroid therapy is itself dangerous with numerous deleterious side effects. There remains a need for the development of specific inhibitors of inflammation, treatments for inflammation and conditions associated with inflammation, regulators of inflammation stimulating cytokines, and kits associated therewith.